Type 1 insulin-dependent diabetes mellitus is caused by an autoimmune attack on the islets of Langerhans. The susceptibility to this disease is determined both genetically and by environmental events. The elucidation of the underlying mechanisms of this disease has been greatly facilitated by the mapping of genes conferring susceptibility or resistance to diabetes in humans. Further, the availability of animal models, including the non-obese diabetic (NOD) mouse has made it possible to perform fine mapping of the corresponding mouse genes. Idd3 is a critical diabetes-susceptibility gene, which has profound effects on development of disease. NOD mice carrying an Idd3 gene from the diabetes-resistant B6 mouse show a substantially reduced incidence of insulitis and diabetes as well as greatly retarded kinetics of disease development. The Idd3 locus has been mapped to a 780 kb region of chromosome 3. The most likely candidate gene in this region is the IL-2 gene. This gene shows several structural differences between alleles that are derived from resistant or susceptible strains of mice. However, definitive evidence for or against a role of IL-2 has not yet been obtained, since it has not been possible until recently to target genes in the NOD genetic background. In this study we will establish gene knock-in technology on the NOD genetic background by using NOD/129 F1 ES (embryonic stem) cells, which we have recently found to be very effective in all the steps of gene targeting. Specifically, to establish or refute a role of IL-2 in T1 DM, we will use gene targeting to replace the IL-2 gene of the NOD mouse with the IL-2 gene of the diabetes-resistant B6 mouse. We hypothesize that the NOD allele of IL-2 confers susceptibility to diabetes in one of two ways. First, IL-2 is required for antigen induced cell death (AICD) and we propose that AICD in NOD mice is deficient as a result of a defective IL-2 gene. Our second hypothesis proposes that suppressor (also called regulatory) T-cells, which are IL-2 dependent, are defective in the NOD mouse as a result of an IL-2 deficiency in this strain. We will utilize our IL-2 knock-in mice to test these two mutually nonexclusive hypotheses.